Enzymatic resolution of selective estrogen receptor modulators

ABSTRACT

The present invention relates to a process for preparing 5-substitued-6-cyclic-5,6,7,8-tetrahydronaphthalen-2-ol compounds useful as an estrogen agonist.

This non-provisional application is based upon and claims priority fromprovisional patent application No. 60/202,418, filed May 8, 2000.

BACKGROUND OF THE INVENTION

The present invention relates to a process for preparing5-substitued-6-cyclic-5,6,7,8-tetrahydronaphthalen-2-ol compounds usefulas an estrogen agonist.

The value of naturally occurring estrogens and synthetic compoundsdemonstrating “estrogenic” activity has been in their medical andtherapeutic uses. A traditional listing of the therapeutic applicationsfor estrogens alone or in combination with other active agents includes:oral contraception; relief for the symptoms of menopause; prevention ofthreatened or habitual abortion; relief of dysmenorrhea; relief ofdysfunctional uterine bleeding; an aid in ovarian development; treatmentof acne; diminution of excessive growth of body hair in women(hirsutism); the prevention of cardiovascular disease; treatment ofosteoporosis; treatment of prostatic carcinoma; and suppression ofpost-partum lactation [Goodman and Gilman, The Pharmacological Basis ofTherapeutics (Seventh Edition) Macmillan Publishing Company, 1985, pages1421-1423]. Accordingly, there has been increasing interest in findingnewly synthesized compounds and new uses for previously known compoundswhich are demonstrably estrogenic, this is, able to mimic the action ofestrogen in estrogen responsive tissue.

From the viewpoint of pharmacologists interested in developing new drugsuseful for the treatment of human diseases and specific pathologicalconditions, it is most important to procure compounds with somedemonstrable estrogen-like function but which are devoid ofproliferative side-effects. Exemplifying this latter view, osteoporosis,a disease in which bone becomes increasingly more fragile, is greatlyameliorated by the use of fully active estrogens; however, due to therecognized increased risk of uterine cancer in patients chronicallytreated with active estrogens, it is not clinically advisable to treatosteoporosis in intact women with fully active estrogens for prolongedperiods. Accordingly, estrogen agonists are the primary interest andfocus.

Osteoporosis is a systemic skeletal disease, characterized by low bonemass and deterioration of bone tissue, with a consequent increase inbone fragility and susceptibility to fracture. In the U.S., thecondition affects more than 25 million people and causes more than 1.3million fractures each year, including 500,000 spine, 250,000 hip and240,000 wrist fractures annually. These cost the nation over $10billion. Hip fractures are the most serious, with 5%-20% of patientsdying within one year, and over 50% of survivors being incapacitated.

The elderly are at greatest risk of osteoporosis, and the problem istherefore predicted to increase significantly with the aging of thepopulation. Worldwide fracture incidence is forecast to increasethree-fold over the next 60 years, and one study estimates that therewill be 4.5 million hip fractures worldwide in 2050.

Women are at greater risk of osteoporosis than men. Women experience asharp acceleration of bone loss during the five years followingmenopause. Other factors that increase risk include smoking, alcoholabuse, a sedentary lifestyle and low calcium intake.

Estrogen is the agent of choice in preventing osteoporosis or postmenopausal bone loss in women; it is the only treatment whichunequivocally reduces fractures. However, estrogen stimulates the uterusand is associated with an increased risk of endometrial cancer. Althoughthe risk of endometrial cancer is thought to be reduced by a concurrentuse of a progestogen, there is still concern about possible increasedrisk of breast cancer with the use of estrogen.

SUMMARY OF THE INVENTION

The present invention relates to a process (Process A) for preparing acompound of the formula

wherein:

A is selected from CH₂ and NR;

B, D and E are independently selected from CH and N;

Y is

(a) phenyl, optionally substituted with 1-3 substituents independentlyselected from R⁴;

(b) naphthyl, optionally substituted with 1-3 substituents independentlyselected from R⁴;

(c) C₃-C₈ cycloalkyl, optionally substituted with 1-2 substituentsindependently selected from R⁴;

(d) C₃-C₈ cycloalkynyl, optionally substituted with 1-2 substituentsindependently selected from R⁴;

(e) a five membered heterocycle containing up to two heteroatomsselected from the group consisting of —O—, —NR²— and —S(O)_(n)—,optionally substituted with 1-3 substituents independently selected fromR⁴;

(f) a six membered heterocycle containing up to two heteroatoms selectedfrom the group consisting of —O—, —NR²— and —S(O)_(n)— optionallysubstituted with 1-3 substituents independently selected from R⁴; or

(g) a bicyclic ring system consisting of a five or six memberedheterocyclic ring fused to a phenyl ring, said heterocyclic ringcontaining up to two heteroatoms selected from the group consisting of—O—, —NR²—, NR²— and —S(O)_(n)—, optionally substituted with 1-3substituents independently selected from R⁴;

Z¹ is

(a) —(CH₂)_(p)W(CH₂)_(q)—;

(b) —O(CH₂)_(p)CR⁵R⁶—;

(c) —O(CH₂)_(p)W(CH₂)_(q);

(d) —OCHR²CHR³; or

(e) —SCHR²CHR³—;

G is

(a) —NR⁷R⁸;

(b)

 wherein n is 0, 1 or 2; m is 1, 2 or 3; Z² is —NH—, —O—, —S—, or —CH₂—;optionally fused on adjacent carbon atoms with one or two phenyl ringsand, optionally independently substituted on carbon with one to threesubstituents and, optionally, independently on nitrogen with achemically suitable substituent selected from R⁴; or

(c) a bicyclic amine containing five to twelve carbon atoms, eitherbridged or fused and optionally substituted with 1-3 substituentsindependently selected from R⁴;

Z¹ and G in combination may be

W is

(a) —CH₂—;

(b) —CH═CH—;

(c) —O—;

(d) —NR²—;

(e) —S(O)_(n)—;

(f)

(g) —CR²(OH)—;

(h) —CONR²—;

(i) —NR²CO—;

(j)

(k) —C≡C—;

R is hydrogen or C₁-C₆ alkyl;

R² and R³ are independently

(a) hydrogen; or

(b) C₁-C₄ alkyl;

R⁴ is

(a) hydrogen;

(b) halogen;

(c) C₁-C₆ alkyl;

(d) C₁-C₄ alkoxy;

(e) C₁-C₄ acyloxy;

(f) C₁-C₄ alkylthio;

(g) C₁-C₄ alkylsulfinyl;

(h) C₁-C₄ alkylsulfonyl;

(i) hydroxy (C₁-C₄)alkyl;

(j) aryl (C₁-C₄)alkyl;

(k) —CO₂H;

(l) —CN;

(m) —CONHOR;

(n) —SO₂NHR;

(o) —NH₂;

(p) C₁-C₄ alkylamino;

(q) C₁-C₄ dialkylamino;

(r) —NHSO₂R;

(s) —NO₂;

(t) -aryl; or

(u) —OH.

R⁵ and R⁶ are independently C₁-C₈ alkyl or together form a C₃-C₁₀carbocyclic ring;

R⁷ and R⁸ are independently

(a) phenyl;

(b) a C₃-C₁₀ carbocyclic ring, saturated or unsaturated;

(c) a C₃-C₁₀ heterocyclic ring containing up to two heteroatoms,selected from —O—, —N— and —S—;

(d) H;

(e) C₁-C₆ alkyl; or

(f) form a 3 to 8 membered nitrogen containing ring with R⁵ or R⁶;

R⁷ and R⁸ in either linear or ring form may optionally be substitutedwith up to three substituents independently selected from C₁-C₆ alkyl,halogen, alkoxy, hydroxy and carboxy;

a ring formed by R⁷ and R⁸ may be optionally fused to a phenyl ring;

e is 0, 1 or 2;

m is 1, 2 or 3;

n is 0, 1 or 2;

p is 0, 1, 2 or 3;

q is 0, 1, 2 or 3;

and optical and geometric isomers thereof;

comprising selectively deacetylating a compound of the formula

wherein R¹ is (C₁-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl wherein thealkyl, alkenyl or alkynyl groups are optionally substituted by one tothree halo, in the presence of a hydrolytic enzyme and an aqueous buffersolution.

The terms “GC-4, PS30, AY30, PGE, AK, N, L-10, AP-12, FAP-15, R-10, G,MAP10 and SAM II” are names of lipase enzymes used in this invention andsold by Amano Chemical Co., 1157 North Main Street, Lombard, Ill. 60148.

The terms “lipase from Pseudomonas fluorescens, lipase from Candidacylindracea, lipase from Mucor miehei, lipase from Wheat germ, lipasefrom Rhizopus arrhizus, lipase from Mucor javanicus, lipase fromPseudomonas cepacia, lipase from Cadia lipolytica and lipase fromPenicillium roqueforfti” are names of lipase enzymes used in thisinvention and sold by Fluka Chemical Co., 1001 West St. Paul Avenue,Milwaukee, Wis. 53233.

The terms “lipoprotein lipase ca#70-6571-01, and lipoprotein lipase ca#70-1481-01” are names of lipase enzymes used in this invention and soldby Genzyme Chemical Co., One Kendall Square, Cambridge, Mass. 02139.

The terms “lipase from Candida cylindracea, lipase from Chromobacteriumviscosum, lipase from Mucor miehei, lipase from Pancreatic, lipase fromPseudomonas fluorescens and lipase from Rhizopus niveus” are names oflipase enzymes used in this invention and sold by RecombinantBiocatalysis Chemical Co., 512 Elmwood Avenue, Sharon Hill, Pa. 19079.

The term “PPL, type II” is a name of a lipase enzyme used in thisinvention and sold by Sigma Chemical Co., P. O. Box 14508, St. Louis,Mo. 63178.

The term “Lip-300″” is a name of a lipase enzyme used in this inventionand sold by TOYOBO Chemical Co., 1450 Broadway, New York, N.Y. 10018.

The terms “immobilized, hog liver, esterase from Hog pancreas, esterasefrom Thernoanaerobium brockii, esterase from Bacillus sp and esterasefrom Mucor miehi” are names of esterase enzymes used in this inventionand sold by Fluka Chemical Co., 1001 West St. Paul Avenue, Milwaukee,Wis. 53233.

The terms “cholesterol esterase from Porcine pancreas, cholesterolesterase from Bovine pancreas, cholesterol esterase from Pseudonomasfluorescens, cholesterol esterase from Porcine liver, cholesterolesterase from Rabbit liver, cholinesterase, cholinesterase from Electriceel, cholinesterase, choloylglycine hydrolase and Porcine liver E-3128”are names of esterase enzymes used in this invention and sold by SigmaChemical Co., P. O. Box 14508, St. Louis, Mo. 63178.

The terms “cholesterin-esterase and cholesterin esterase fromPseudonomas fluorescents” are names of esterase enzymes used in thisinvention and sold by Boehringer Mannheim Chemical Co., 9115 Hague Road,Indianapolis, Ind. 46250.

The terms “cat I-1256, dog I7379, eel I-1266, horse I9627, calf I7876,guinea pig I1631, mouse I8254, goat I2635, chicken I8001, sheep I0132,pigeon I8376, seal I7627, rattlesnake I9885, trout I5131, turtle I-0757,rat I1380, lungfish I7377, salmon I7502, eel (electrophorus electricus)I8380 and lemon shark I1130” are names of liver acetone powder enzymesused in this invention and sold by Sigma Chemical Co., P.O. Box 14508,St. Louis, Mo. 63178.

The present invention further relates to Process A wherein thehydrolytic enzyme is lipase.

The present invention further relates to Process A wherein thehydrolytic enzyme is esterase.

The present invention further relates to Process A wherein thehydrolytic enzyme is liver acetone powder.

The present invention further relates to Process A wherein the lipase isGC-4, PS30, AY30, PGE, AK, N, L-10, AP-12, FAP-15, R-10, G, MAP10, SAMII, lipase from Pseudomonas fluorescens, lipase from Candidacylindracea, Lip-300, lipase from Chromobacterium viscosum, lipase fromMucor miehei, lipase from Pancreatic, Pseudomonas fluorescens, lipasefrom Rhizopus niveus, PPL, type II, lipase from Wheat germ, lipase fromRhizopus arrhizus, lipase from Mucor javanicus, lipase from Pseudomonascepacia, lipase from Cadia lipolytica, lipase from Penicilliumroqueforti, lipoprotein lipase ca#70-6571-01, lipase from Porcinepancreas and lipoprotein lipase ca# 70-1481-01.

The present invention further relates to Process A wherein the esteraseis PLE-A, immobilized, hog liver, esterase from Hog pancreas, Porcineliver E-3128, cholesterin-esterase, cholesterol esterase fromPseudonomas fluorescens, cholesterol esterase from Porcine pancreas,cholesterol esterase from Bovine pancreas, cholesterol esterase fromPseudonomas fluorescens, cholesterol esterase from Porcine liver,cholesterol esterase from Rabbit liver, cholinesterase, cholinesterasefrom Electric eel, cholinesterase, choloylglycine hydrolase, esterasefrom Thermoanaerobium brockii, esterase from Bacillus sp and esterasefrom Mucor miehi.

The present invention further relates to Process A wherein the liveracetone powder is cat I-1256, dog I7379, eel I-1266, horse I9627, calfI7876, guinea pig I1631, mouse I8254, goat I2635, chicken I8001, sheepI0132, pigeon I8376, seal I7627, rattlesnake I9885, trout I5131, turtleI-0757, rat I1380, lungfish I7377, salmon I7502, eel (electrophoruselectricus) I8380 and lemon shark I1130.

The present invention further relates to Process A wherein thehydrolytic enzyme is immobilized on a solid support.

The present invention further relates to Process A wherein thehydrolytic enzyme is in pure crystalline form.

The present invention further relates to Process A wherein the aqueousbuffer solution is a phosphate, citric acid or boronic acid solution.

The present invention further relates to Process A wherein the aqueousbuffer solution has a pH between a pH of about 6 to a pH of about 8.

The present invention relates to a process for preparing a compound ofthe formula

comprising selectively deacetylating a compound of the formula

wherein R¹ is (C₁-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl wherein thealkyl, alkenyl or alkynyl groups are optionally substituted by one tothree halo in the presence of a hydrolytic enzyme and an aqueous buffersolution.

The present invention relates to a process (Process B) for preparing acompound of the formula:

wherein:

A is selected from CH₂ and NR;

B, D and E are independently selected from CH and N;

Y is

(a) phenyl, optionally substituted with 1-3 substituents independentlyselected from R⁴;

(b) naphthyl, optionally substituted with 1-3 substituents independentlyselected from R⁴;

(c) C₃-C₈ cycloalkyl, optionally substituted with 1-2 substituentsindependently selected from R⁴;

(d) C₃-C₈ cycloalkynyl, optionally substituted with 1-2 substituentsindependently selected from R⁴;

(e) a five membered heterocycle containing up to two heteroatomsselected from the group consisting of —O—, —NR²— and —S(O)_(n)—,optionally substituted with 1-3 substituents independently selected fromR⁴;

(f) a six membered heterocycle containing up to two heteroatoms selectedfrom the group consisting of —O—, —NR²— and —S(O)_(n)— optionallysubstituted with 1-3 substituents independently selected from R⁴; or

(g) a bicyclic ring system consisting of a five or six memberedheterocyclic ring fused to a phenyl ring, said heterocyclic ringcontaining up to two heteroatoms selected from the group consisting of—O—, —NR²—, NR²— and —S(O)_(n)—, optionally substituted with 1-3substituents independently selected from R⁴;

Z¹ is

(a) —(CH₂)_(p)W(CH₂)_(q)—;

(b) —O(CH₂)_(p)CR⁵R⁶—;

(c) —O(CH₂)_(p)W(CH₂)_(q);

(d) —OCHR²CHR³—; or

(e) —SCHR²CHR³—;

G is

(a) —NR⁷R⁸;

(b)

 wherein n is 0, 1 or 2; m is 1, 2 or 3; Z² is —NH—, —O—, —S—, or —CH₂—;optionally fused on adjacent carbon atoms with one or two phenyl ringsand, optionally independently substituted on carbon with one to threesubstituents and, optionally, independently on nitrogen with achemically suitable substituent selected from R⁴; or

(c) a bicyclic amine containing five to twelve carbon atoms, eitherbridged or fused and optionally substituted with 1-3 substituentsindependently selected from R⁴;

Z¹ and G in combination may be

W is

(a) —CH₂—;

(b) —CH═CH—;

(c) —O—;

(d) —NR²—;

(e) —S(O)_(n)—;

(f)

(g) —CR²(OH)—;

(h) —CONR²—;

(i) —NR²CO—;

(j)

(k) —C≡C—;

R is hydrogen or C₁-C₆ alkyl;

R² and R³ are independently

(a) hydrogen; or

(b) C₁-C₄ alkyl;

R⁴ is

(a) hydrogen;

(b) halogen;

(c) C₁-C₆ alkyl;

(d) C₁-C₄ alkoxy;

(e) C₁-C₄ acyloxy;

(f) C₁-C₄ alkylthio;

(g) C₁-C₄ alkylsulfinyl;

(h) C₁-C₄ alkylsulfonyl;

(i) hydroxy (C₁-C₄)alkyl;

(j) aryl (C₁-C₄)alkyl;

(k) —CO₂H;

(l) —CN;

(m) —CONHOR;

(n) —SO₂NHR;

(o) —NH₂;

(p) C₁-C₄ alkylamino;

(q) C₁-C₄ dialkylamino;

(r) —NHSO₂R;

(s) —NO₂;

(t) -aryl; or

(u) —OH.

R⁵ and R⁶ are independently C₁-C₈ alkyl or together form a C₃-C₁₀carbocyclic ring;

R⁷ and R⁸ are independently

(a) phenyl;

(b) a C₃-C₁₀ carbocyclic ring, saturated or unsaturated;

(c) a C₃-C₁₀ heterocyclic ring containing up to two heteroatoms,selected from —O—, —N— and —S—;

(d) H;

(e) C₁-C₆ alkyl; or

(f) form a 3 to 8 membered nitrogen containing ring with R⁵ or R⁰;

R⁷ and R⁸ in either linear or ring form may optionally be substitutedwith up to three substituents independently selected from C₁-C₆ alkyl,halogen, alkoxy, hydroxy and carboxy;

a ring formed by R⁷ and R⁸ may be optionally fused to a phenyl ring;

e is 0, 1 or 2;

m is 1, 2 or 3;

n is 0, 1 or 2;

p is 0, 1, 2 or 3;

q is 0, 1, 2 or 3;

and optical and geometric isomers thereof;

comprising enzymatically resolving of a compound of the formula

wherein R¹ is (C₁-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl wherein thealkyl, alkenyl or alkynyl groups are optionally substituted by one tothree halo in the presence of a lipase and an aqueous buffer solution;and (b) reacting the compound of formula IV so formed

wherein R¹ is as defined above, with a base in the presence of a polarprotic solvent.

The present invention further relates to Process B wherein the aqueousbuffer solution is a phosphate, citric acid or boronic acid solution.

The present invention further relates to Process B wherein the lipasefrom Mucor miehei.

The present invention further relates to Process B wherein the base issodium methoxy, sodium hydroxide, lithium hydroxide or potassiumhydroxide.

The present invention further relates to Process B wherein the polarprotic solvent is methanol, ethanol or water.

The present invention further relates to Process B wherein the lipase isimmobilized on a solid support.

The present invention further relates to Process B wherein the lipase isa cross-linked enzyme.

The present invention further relates to Process B wherein the lipase isin pure crystalline form.

The present invention relates to a process for preparing a compound ofthe formula

comprising enzymatically resolving of a compound of the formula

wherein R¹ is (C₁-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl wherein thealkyl, alkenyl or alkynyl groups are optionally substituted by one tothree halo in the presence of a lipase and an aqueous buffer solution;and (b) reacting the compound of Formula X so formed

wherein R¹ is as defined above, with a base in the presence of a polarprotic solvent.

The present invention relates to a process (Process C) for preparing acompound of the formula:

wherein:

A is selected from CH₂ and NR;

B, D and E are independently selected from CH and N;

Y is

(a) phenyl, optionally substituted with 1-3 substituents independentlyselected from R⁴;

(b) naphthyl, optionally substituted with 1-3 substituents independentlyselected from R⁴;

(c) C₃-C₈ cycloalkyl, optionally substituted with 1-2 substituentsindependently selected from R⁴;

(d) C₃-C₈ cycloalkynyl, optionally substituted with 1-2 substituentsindependently selected from R⁴;

(e) a five membered heterocycle containing up to two heteroatomsselected from the group consisting of —O—, —NR²— and —S(O)_(n)—,optionally substituted with 1-3 substituents independently selected fromR⁴;

(f) a six membered heterocycle containing up to two heteroatoms selectedfrom the group consisting of —O—, —NR²— and —S(O)_(n)— optionallysubstituted with 1-3 substituents independently selected from R⁴; or

(g) a bicyclic ring system consisting of a five or six memberedheterocyclic ring fused to a phenyl ring, said heterocyclic ringcontaining up to two heteroatoms selected from the group consisting of—O—, —NR²—, NR²— and —S(O)_(n)—, optionally substituted with 1-3substituents independently selected from R⁴;

Z¹ is

(a) —(CH₂)_(p)W(CH₂)_(q)—;

(b) —O(CH₂)_(p)CR⁵R⁸—;

(c) —O(CH₂)_(p)W(CH₂)_(q);

(d) —OCHR²CHR³; or

(e) —SCHR²CHR³—;

G is

(a) —NR⁷R⁸;

(b)

 wherein n is 0, 1 or 2; m is 1, 2 or 3; Z² is —NH—, —O—, —S—, or —CH₂—;optionally fused on adjacent carbon atoms with one or two phenyl ringsand, optionally independently substituted on carbon with one to threesubstituents and, optionally, independently on nitrogen with achemically suitable substituent selected from R⁴; or

(c) a bicyclic amine containing five to twelve carbon atoms, eitherbridged or fused and optionally substituted with 1-3 substituentsindependently selected from R⁴;

Z¹ and G in combination may be

W is

(a) —CH₂—;

(b) —CH═CH—;

(c) —O—;

(d) —NR²;

(e) —S(O)_(n)—;

(f)

(g) —CR²(OH)—;

(h) —CONR²—;

(j) —NR²CO—;

(k) —C≡C—;

R is hydrogen or C₁-C₆ alkyl;

R² and R³ are independently

(a) hydrogen; or

(b) C₁-C₄ alkyl;

R⁴ is

(a) hydrogen;

(b) halogen;

(c) C₁-C₆ alkyl;

(d) C₁-C₄ alkoxy;

(e) C₁-C₄ acyloxy;

(f) C₁-C₄ alkylthio;

(g) C₁-C₄ alkylsulfinyl;

(h) C₁-C₄ alkylsulfonyl;

(i) hydroxy (C₁-C₄)alkyl;

(j) aryl (C₁-C₄)alkyl;

(k) —CO₂H;

(l) —CN;

(m) —CONHOR;

(n) —SO₂NHR;

(o) —NH₂;

(p) C₁-C₄ alkylamino;

(q) C₁-C₄ dialkylamino;

(r) —NHSO₂R;

(s) —NO₂;

(t) -aryl; or

(u) —OH.

R⁵ and R⁶ are independently C₁-C₈ alkyl or together form a C₃-C₁₀carbocyclic ring;

R⁷ and R⁸ are independently

(a) phenyl;

(b) a C₃-C₁₀ carbocyclic ring, saturated or unsaturated;

(c) a C₃-C₁₀ heterocyclic ring containing up to two heteroatoms,selected from —O—, —N— and —S—;

(d) H;

(e) C₁-C₆ alkyl; or

(f) form a 3 to 8 membered nitrogen containing ring with R⁵ or R⁶;

R⁷ and R⁸ in either linear or ring form may optionally be substitutedwith up to three substituents independently selected from C₁-C₆ alkyl,halogen, alkoxy, hydroxy and carboxy;

a ring formed by R⁷ and R⁸ may be optionally fused to a phenyl ring;

e is 0, 1 or 2;

m is 1, 2 or 3;

n is 0, 1 or 2;

p is 0, 1, 2 or 3;

q is 0, 1, 2 or 3;

and optical and geometric isomers thereof;

comprising enzymatically resolving of a compound of the formula

in the presence of a lipase and an acetylating agent, and (b) reactingthe compound of formula IV so formed

wherein R¹ is (C₁-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl wherein thealkyl, alkenyl or alkynyl groups are optionally substituted by one tothree halo, with a base in the presence of a polar protic solvent.

The present invention further relates to Process C wherein thehydrolytic enzyme is a lipase.

The present invention further relates to Process C wherein the lipase isGC-4, PS30, AY30, PGE, AK, N, L-10, AP-12, FAP-15, R-10, G, MAP10, SAMII, lipase from Pseudomonas fluorescens, lipase from Candidacylindracea, Lip-300, lipase from Chromobacterium viscosum, lipase fromMucor miehei, lipase from Pancreatic, lipase from Pseudomonasfluorescens, lipase from Rhizopus niveus, PPL, type II, lipase fromWheat germ, lipase from Rhizopus arrhizus, lipase from Mucor javanicus,lipase from Pseudomonas cepacia, lipase from Cadia lipolytica, lipasefrom Penicillium roqueforti, lipoprotein lipase ca #70-6571-01, lipasefrom Porcine pancreas, and lipoprotein lipase ca #70-1481-01.

The present invention further relates to Process C wherein theacetylating agent is ethyl acetate, vinyl acetate, chloroacetate ortrifluoroacetate.

The present invention further relates to Process C wherein the base issodium methoxy, sodium hydroxide, lithium hydroxide or potassiumhydroxide.

The present invention further relates to Process C wherein the polarprotic solvent is methanol, ethanol or water.

The present invention further relates to Process C wherein the lipase isimmobilized on a solid support.

The present invention further relates to Process C wherein the lipase isa cross-linked enzyme.

The present invention further relates to Process C wherein the lipase isin pure crystalline form.

The present invention relates to a process for preparing a compound ofthe formula

comprising enzymatically resolving of a compound of the formula

in the presence of a lipase and acetylating agent, and (b) reacting thecompound of Formula X so formed

wherein R¹ is (C₁-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl wherein thealkyl, alkenyl or alkynyl groups are optionally substituted by one tothree halo, with a base in the presence of a polar protic solvent.

The present invention relates to a process (Process D) for preparing acompound of the formula:

wherein:

A is selected from CH₂ and NR;

B, D and E are independently selected from CH and N;

Y is

(a) phenyl, optionally substituted with 1-3 substituents independentlyselected from R⁴;

(b) naphthyl, optionally substituted with 1-3 substituents independentlyselected from R⁴;

(c) C₃-C₈ cycloalkyl, optionally substituted with 1-2 substituentsindependently selected from R⁴;

(d) C₃-C₈ cycloalkynyl, optionally substituted with 1-2 substituentsindependently selected from R⁴;

(e) a five membered heterocycle containing up to two heteroatomsselected from the group consisting of —O—, —NR²— and —S(O)_(n)—,optionally substituted with 1-3 substituents independently selected fromR⁴;

(f) a six membered heterocycle containing up to two heteroatoms selectedfrom the group consisting of —O—, —NR²— and —S(O)_(n)— optionallysubstituted with 1-3 substituents independently selected from R⁴; or

(g) a bicyclic ring system consisting of a five or six memberedheterocyclic ring fused to a phenyl ring, said heterocyclic ringcontaining up to two heteroatoms selected from the group consisting of—O—, —NR²—, NR²— and —S(O)_(n)—, optionally substituted with 1-3substituents independently selected from R⁴;

Z¹ is

(a) —(CH₂)_(p)W(CH₂)_(q)—;

(b) —O(CH₂)_(p)CR⁵R⁶—;

(c) —O(CH₂)_(p)W(CH₂)_(q);

(d) —OCHR²CHR³—; or

(e) —SCHR²CHR³—;

G is

(a) —NR⁷R⁸;

(b)

 wherein n is 0, 1 or 2; m is 1, 2 or 3; Z² is —NH—, —O—, —S—, or —CH₂—;optionally fused on adjacent carbon atoms with one or two phenyl ringsand, optionally independently substituted on carbon with one to threesubstituents and, optionally, independently on nitrogen with achemically suitable substituent selected from R⁴; or

(c) a bicyclic amine containing five to twelve carbon atoms, eitherbridged or fused and optionally substituted with 1-3 substituentsindependently selected from R⁴;

Z¹ and G in combination may be

W is

(a) —CH₂—;

(b) —CH═CH—;

(c) —O—;

(d) —NR²—;

(e) —S(O)_(n)—;

(f)

(g) —CR²(OH)—;

(h) —CONR²—;

(i) —NR²CO—;

(j)

(k) —C≡C—;

R is hydrogen or C₁-C₆ alkyl;

R² and R³ are independently

(a) hydrogen; or

(b) C₁-C₄ alkyl;

R⁴ is

(a) hydrogen;

(b) halogen;

(c) C₁-C₆ alkyl;

(d) C₁-C₄ alkoxy;

(e) C₁-C₄ acyloxy;

(f) C₁-C₄ alkylthio;

(g) C₁-C₄ alkylsulfinyl;

(h) C₁-C₄ alkylsulfonyl;

(i) hydroxy (C₁-C₄)alkyl;

(j) aryl (C₁-C₄)alkyl;

(k) —CO₂H;

(l) —CN;

(m) —CONHOR;

(n) —SO₂NHR;

(o) —NH₂;

(p) C₁-C₄ alkylamino;

(q) C₁-C₄ dialkylamino;

(r) —NHSO₂R;

(s) —NO₂;

(t) -aryl; or

(u) —OH.

R⁵ and R⁶ are independently C₁-C₈ alkyl or together form a C₃-C₁₀carbocyclic ring;

R⁷ and R⁸ are independently

(a) phenyl;

(b) a C₃-C₁₀ carbocyclic ring, saturated or unsaturated;

(c) a C₃-C₁₀ heterocyclic ring containing up to two heteroatoms,selected from —O—, —N— and —S—;

(d) H;

(e) C₁-C₆ alkyl; or

(f) form a 3 to 8 membered nitrogen containing ring with R⁵ or R⁶;

R⁷ and R⁸ in either linear or ring form may optionally be substitutedwith up to three substituents independently selected from C₁-C₆ alkyl,halogen, alkoxy, hydroxy and carboxy;

a ring formed by R⁷ and R⁸ may be optionally fused to a phenyl ring;

e is 0, 1 or 2;

m is 1, 2 or 3;

n is 0, 1 or 2;

p is 0, 1, 2 or 3;

q is 0, 1, 2 or 3;

and optical and geometric isomers thereof;

comprising enzymatically resolving of a compound of the formula

in the presence of lipase.

The present invention further relates to Process D wherein the lipase isMucor miehei.

The present invention further relates to Process D wherein the lipase isimmobilized on a solid support.

The present invention further relates to Process D wherein the lipase isa cross-linked enzyme.

The present invention further relates to Process D wherein the lipase isin pure crystalline form.

The present invention relates to a process for preparing a compound ofthe formula

comprising enzymatically resolving of a compound of the formula

in the presence of lipase.

The starting materials for the present invention are prepared accordingto U.S. Pat. No. 5,552,412, which is incorporated by reference in itsentirety.

In reaction 1 of Scheme 1, the compound of formula II is converted tothe corresponding compounds of formulas I and III by enzymaticresolution of II in the presence of a hydrolytic enzyme, mobilized orimmobilized on different solid support, a cross-linked enzyme or acrystallized enzyme. Hydrolytic enzymes include (a) lipase (GC-4, PS30,AY30, PGE, AK, N, L-10, AP-12, FAP-15, R-10, G, MAP10, SAM II, lipasefrom Pseudomonas fluorescens, lipase from Candida cylindracea, Lip-300,lipase from Chromobacterium viscosum, lipase from Mucor miehei, lipasefrom Pancreatic, lipase from Pseudomonas fluorescens, lipase fromRhizopus niveus, PPL, type II, lipase from Wheat germ, lipase fromRhizopus arrhizus, lipase from Mucor javanicus, lipase from Pseudomonascepacia, lipase from Cadia lipolytica, lipase from Penicilliumroqueforti, lipoprotein lipase ca #70-6571-01, lipase from Porcinepancreas, and lipoprotein lipase ca #70-1481-01), (b) esterase (PLE-A,immobilized, hog liver, esterase from Hog pancreas, Porcine liverE-3128, cholesterin-esterase, cholesterol esterase from Pseudonomasfluorescens, cholesterol esterase from Porcine pancreas, cholesterolesterase from Bovine pancreas, cholesterol esterase from Pseudonomasfluorescens, cholesterol esterase from Porcine liver, cholesterolesterase from Rabbit liver, cholinesterase, cholinesterase from Electriceel, cholinesterase, choloylglycine hydrolase, esterase fromThermoanaerobium brockii, esterase from Bacillus sp and esterase fromMucor miehei) or (c) liver acetone powder (cat I-1256, dog I7379, eelI-1266, horse I9627, calf I7876, guinea pig I1631, mouse I8254, goatI2635, chicken I8001, sheep I0132, pigeon I8376, seal I7627, rattlesnakeI9885, trout I5131, turtle I-0757, rat I1380, lungfish I7377, salmonI7502, eel (electrophorus electricus) I8380 and lemon shark I1130). Thereaction is carried out in an aqueous buffer solution, such as aphosphate, citric acid or boronic acid buffer, with or without anorganic solvent, such as methylene chloride, tetrahydrofuran, acetone,dimethyl formamide or dioxane. The buffer solution has a pH of about 6to a pH of about 8, preferably a pH of about 7. The reaction is stirredat a temperature between room temperature to about 65° C., preferablyabout 25° C. to about 40° C., for a time period dependent upon theenzyme employed and the desired enantiomeric excess conversion of theracemic mixture.

In reaction I of Scheme 2, the compound of formula II is converted tothe corresponding compounds of formulas IV and V by enzymatic resolutionof II with a lipase, such as Mucor miehei, according to the proceduredescribed above in reaction I of Scheme 1.

In reaction I of Scheme 3, the compound of formula VI is converted tothe corresponding compounds of formulas IV and V by enzymatic resolutionof VI with a lipase (GC-4, PS30, AY30, PGE, AK, N, L-10, AP-12, FAP-15,R-10, G, MAP10, SAM II, lipase from Pseudomonas fluorescens, lipase fromcylindracea, Lip-300, lipase from Candida cylindracea, lipase fromChromobacterium viscosum, lipase from Mucor miehei, lipase fromPancreatic, lipase from Pseudomonas fluorescens, lipase from Rhizopusniveus, PPL, type II, lipase from Wheat germ, lipase from Rhizopusarrhizus, lipase from Mucor javanicus, lipase from Pseudomonas cepacia,lipase from Cadia lipolytica, lipase from Penicillium roqueforti,lipoprotein lipase ca #70-6571-01, lipase from Porcine pancreas, andlipoprotein lipase ca #70-1481-01) in the presence of an acetylatingagent, such as ethyl acetate, vinyl acetate, chloroacetate ortrifluoroacetate. The reaction is stirred in an aprotic solvent, such asmethylene chlorine, ethylene glycol, dimethyl ether, dichloroethane,hexane, tetrahydrofuran and dioxane, at a temperature between roomtemperature to about 65° C., preferably about 25° C. to about 40° C.,for a time period dependent upon the enzyme employed and the desiredenantiomeric excess conversion of the racemic mixture.

In reaction I of Scheme 4, the compound of formula VI is converted tothe corresponding compounds of formulas of I and III by enzymaticresolution of II with a lipase, such as Mucor miehei. The reaction isstirred in an aprotic solvent, such as methylene chlorine, ethyleneglycol, dimethyl ether, dichloroethane, hexane, tetrahydrofuran anddioxane, at a temperature between room temperature to about 65° C.,preferably about 25° C. to about 40° C., for a time period dependantupon the enzyme employed and the desired enantiomeric excess conversionof the racemic mixture.

In reaction I of Scheme 5, the compound of formula IV is converted tothe corresponding compound of formula I by deacetylating IV with a base,such as sodium methoxy, sodium hydroxide, lithium hydroxide or potassiumhydroxide, in a polar protic solvent, such as methanol ethanol or water.The reaction is stirred at room temperature for a time period betweenabout 1 hour and about 24 hours, preferably about 6 hours.

EXAMPLE 1(−)Cis-6(S)-Phenyl-5(R)-[4-(2-pyrrolidin-1-yl-ethoxy)Phenyl]-5,6,7,8-tetrahydronaphthalen-2-ol

A mixture of the racemic acetylate, corresponding to the title compound,cholesterol esterase from Porcine pancreas (C-9530), commercially soldby SIGMA, and a 0.1M phosphate buffer, having a pH of 7, was stirred atroom temperature. The enzymatic resolution was stopped at 30 to 50%conversion and monitored by high pressure liquid chromatography. Thesolution was then extracted with ethyl acetate. The organic layer waswashed with water followed by brine, dried over magnesium sulfate,filtered and concentrated to yield a brown solid. The residue wasdissolved in methylene chloride and purified by flash chromatographyover SiO₂ with methylene chloride/methanol in a 9:1 ratio and a fewdrops of NH₄OH as the elutant to yield the product as an off-white foam.¹H NMR (250M Hz, CDCL₃): d 7.04 (m, 3H), 6.74 (m, 2H), 6.63 (d, J=8.3Hz, 2H), 6.50 (m, 3H), 6.28 (d, J=8.6 Hz, 2H), 4.14 (d, J=4.9 Hz, 1H),3.94 (t, J=5.3 Hz, 2H), 3.249dd, J=12.5, 4.1 Hz, 1H), 2.59 (m, 4H), 2.78(m, 1H), 1.88 (m, 4H), 1.68 (m, 1H). The optical purity of the productwas determined by chiral HPLC assay.

What is claimed is:
 1. A process for preparing a compound of theformula:

wherein: A is selected from CH₂ and NR; B, D and E are independentlyselected from CH and N; Y is (a) phenyl, optionally substituted with 1-3substituents independently selected from R⁴; (b) naphthyl, optionallysubstituted with 1-3 substituents independently selected from R⁴; (c)C₃-C₈ cycloalkyl, optionally substituted with 1-2 substituentsindependently selected from R⁴; (d) C₃-C₈ cycloalkynyl, optionallysubstituted with 1-2 substituents independently selected from R⁴; (e) afive membered heterocycle containing up to two heteroatoms selected fromthe group consisting of —O—, —NR²— and —S(O)_(n)—, optionallysubstituted with 1-3 substituents independently selected from R⁴; (f) asix membered heterocycle containing up to two heteroatoms selected fromthe group consisting of —O—, —NR²— and —S(O)_(n)— optionally substitutedwith 1-3 substituents independently selected from R⁴; or (g) a bicyclicring system consisting of a five or six membered heterocyclic ring fusedto a phenyl ring, said heterocyclic ring containing up to twoheteroatoms selected from the group consisting of —O—, —NR²—, NR²— and—S(O)_(n)—, optionally substituted with 1-3 substituents independentlyselected from R⁴; Z¹ is (a) —(CH₂)_(p)W(CH₂)_(q)—; (b)—O(CH₂)_(p)CR⁵R⁶—; (c) —O(CH₂)_(p)W(CH₂)_(q); (d) —OCHR²CHR³—; or (e)—SCHR²CHR³—; G is (a) —NR⁷R⁸; (b)

 wherein n is 0, 1 or 2; m is 1, 2 or 3; Z² is —NH—, —O—, —S—, or —CH₂—;optionally fused on adjacent carbon atoms with one or two phenyl ringsand, optionally independently substituted on carbon with one to threesubstituents and, optionally, independently on nitrogen with achemically suitable substituent selected from R⁴; or (c) a bicyclicamine containing five to twelve carbon atoms, either bridged or fusedand optionally substituted with 1-3 substituents independently selectedfrom R⁴; Z¹ and G in combination may be

W is (a) —CH₂—; (b) —CH═CH—; (c) —O—; (d) —NR²; (e) —S(O)_(n)—; (f)

(g) —CR²(OH)—; (h) —CONR²—; (i) —NR²CO—; (j)

(k) —C≡C—; R is hydrogen or C₁-C₆ alkyl; R² and R³ are independently (a)hydrogen; or (b) C₁-C₄ alkyl; R⁴ is (a) hydrogen; (b) halogen; (c) C₁-C₆alkyl; (d) C₁-C₄ alkoxy; (e) C₁-C₄ acyloxy; (f) C₁-C₄ alkylthio; (g)C₁-C₄ alkylsulfinyl; (h) C₁-C₄ alkylsulfonyl; (i) hydroxy (C₁-C₄)alkyl;(j) aryl (C₁-C₄)alkyl; (k) —CO₂H; (l) —CN; (m) —CONHOR; (n) —SO₂NHR; (o)—NH₂; (p) C₁-C₄ alkylamino; (q) C₁-C₄ dialkylamino; (r) —NHSO₂R; (s)—NO₂; (t) -aryl; or (u) —OH; R⁵ and R⁶ are independently C₁-C₈ alkyl ortogether form a C₃-C₁₀ carbocyclic ring; R⁷ and R⁸ are independently (a)phenyl; (b) a C₃-C₁₀ carbocyclic ring, saturated or unsaturated; (c) aC₃-C₁₀ heterocyclic ring containing up to two heteroatoms, selected from—O—, —N— and —S—; (d) H; (e) C₁-C₆ alkyl; or (f) form a 3 to 8 memberednitrogen containing ring with R⁵ or R⁶; R⁷ and R⁸ in either linear orring form may optionally be substituted with up to three substituentsindependently selected from C₁-C₆ alkyl, halogen, alkoxy, hydroxy andcarboxy; a ring formed by R⁷ and R⁸ may be optionally fused to a phenylring; e is 0, 1 or 2; m is 1, 2 or 3; n is 0, 1 or 2; p is 0, 1, 2 or 3;q is 0, 1, 2 or 3; and optical and geometric isomers thereof; comprisingselectively deacetylating a compound of the formula

wherein R¹ is (C₁-C₆)alkyl, (C₂-C₆)alkenyl, (C₂-C₆)alkynyl wherein thealkyl, alkenyl or alkynyl groups are optionally substituted by one tothree halo, in the presence of an aqueous buffer solution and ahydrolytic enzyme selected from the group consisting of lipase,esterase, and liver acetone powder.
 2. A process according to claim 1,wherein the hydrolytic enzyme is lipase.
 3. A process according to claim1, wherein the hydrolytic enzyme is esterase.
 4. A process according toclaim 1, wherein the hydrolytic enzyme is liver acetone powder.
 5. Aprocess according to claim 1, wherein the hydrolytic enzyme is lipasefrom Porcine pancreas, cholesterol esterase from Pseudomonas Fluorscensand cholesterol esterase from Porcine pancreas.
 6. A process accordingto claim 2, wherein the lipase is GC-4, PS30, AY30, PGE, AK, N, L-10,AP-12, FAP-15, R-10, G, MAP10, SAM II, lipase from Pseudomonasfluorescens, lipase from Candida cylindracea, Lip-300, lipase fromChromobacterium viscosum, lipase from Mucor miehei, lipase fromPancreatic, lipase from Pseudomonas fluorescens, lipase from Rhizopusniveus, PPL, type II, lipase from Wheat germ, lipase from Rhizopusarrhizus, lipase from Mucor javanicus, lipase from Pseudomonas cepacia,lipase from Cadia lipolytica, lipase from Penicillium roqueforti,lipoprotein lipase ca #70-6571-01, lipase from Porcine pancreas, andlipoprotein lipase ca #70-1481-01.
 7. A process according to claim 3,wherein the esterase is PLE-A, immobilized, hog liver, esterase from Hogpancreas, Porcine liver E-3128, cholesterin-esterase, cholesterolesterase from Pseudomonas fluorescens, cholesterol esterase from Porcinepancreas, cholesterol esterase from Bovine pancreas, cholesterolesterase from Pseudonomas fluorescens, cholesterol esterase from Porcineliver, cholesterol esterase from Rabbit liver, cholinesterase,cholinesterase from Electric eel, cholinesterase, choloylglycinehydrolase, esterase from Thermoanaerobium brockii, esterase fromBacillus sp and esterase from Mucor miehi.
 8. A process according toclaim 4, wherein the liver acetone powder is cat I-1256, dog I7379, eelI-1266, horse I9627, calf I7876, guinea pig I1631, mouse I8254, goatI2635, chicken I8001, sheep I0132, pigeon I8376, seal I7627, rattlesnakeI9885, trout I5131, turtle I-0757, rat I1380, lungfish I7377, salmonI7502, eel (electrophorus electricus) I8380 and lemon shark I1130.
 9. Aprocess according to claim 1, wherein the hydrolytic enzyme isimmobilized on a solid support.
 10. A process according to claim 1,wherein the hydrolytic enzyme is a cross-linked enzyme.
 11. A processaccording to claim 1, wherein the lipase is in pure crystalline form.12. A process according to claim 1, wherein the aqueous buffer solutionis a phosphate, citric acid or boronic acid solution.
 13. A processaccording to claim 1, wherein the aqueous buffer solution has a pHbetween a pH of about 6 to a pH of about
 8. 14. A process according toclaim 1, for preparing a compound of the formula

comprising selectively deacetylating a compound of the formula

wherein R¹ is (C₁-C₆)alkyl, (C₂-C₆)alkynyl, (C₂-C₆)alkenyl wherein thealkyl, alkenyl or alkynyl groups are optionally substituted by one tothree halo in the presence of an aqueous buffer solution and ahydrolytic enzyme selected from the group consisting of lipase,esterase, and liver acetone powder.